Streptomyces
Over the last century, Streptomyces bacteria – and their metabolic products – have revolutionized modern medicine. These little pharmaceutical factories produce a vast array of natural products that have been co-opted for medical and agricultural therapies. In addition to their metabolic sophistication, Streptomyces also exhibit remarkable developmental and regulatory complexity.
Guest-edited by Dr Marie Elliot, this collection of keynote research articles will highlight fascinating aspects of Streptomyces biology, and the advances that are providing us with newfound insight and appreciation for these extraordinary bacteria.
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21 - 40 of 44 results
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Developmental delay in a Streptomyces venezuelae glgE null mutant is associated with the accumulation of α-maltose 1-phosphate
More LessThe GlgE pathway is thought to be responsible for the conversion of trehalose into a glycogen-like α-glucan polymer in bacteria. Trehalose is first converted to maltose, which is phosphorylated by maltose kinase Pep2 to give α-maltose 1-phosphate. This is the donor substrate of the maltosyl transferase GlgE that is known to extend α-1,4-linked maltooligosaccharides, which are thought to be branched with α-1,6 linkages. The genome of Streptomyces venezuelae contains all the genes coding for the GlgE pathway enzymes but none of those of related pathways, including glgC and glgA of the glycogen pathway. This provides an opportunity to study the GlgE pathway in isolation. The genes of the GlgE pathway were upregulated at the onset of sporulation, consistent with the known timing of α-glucan deposition. A constructed ΔglgE null mutant strain was viable but showed a delayed developmental phenotype when grown on maltose, giving less cell mass and delayed sporulation. Pre-spore cells and spores of the mutant were frequently double the length of those of the wild-type, implying impaired cross-wall formation, and spores showed reduced tolerance to stress. The mutant accumulated α-maltose 1-phosphate and maltose but no α-glucan. Therefore, the GlgE pathway is necessary and sufficient for polymer biosynthesis. Growth of the ΔglgE mutant on galactose and that of a Δpep2 mutant on maltose were analysed. In both cases, neither accumulation of α-maltose 1-phosphate/α-glucan nor a developmental delay was observed. Thus, high levels of α-maltose 1-phosphate are responsible for the developmental phenotype of the ΔglgE mutant, rather than the lack of α-glucan.
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Isolation and antimicrobial activities of actinobacteria closely associated with liquorice plants Glycyrrhiza glabra L. and Glycyrrhiza inflate BAT. in Xinjiang, China
A total of 218 actinobacteria strains were isolated from wild perennial liquorice plants Glycyrrhiza glabra L. and Glycyrrhiza. inflate BAT. Based on morphological characteristics, 45 and 32 strains from G. inflate and G. glabra, respectively, were selected for further analyses. According to 16S rRNA sequence analysis, most of the strains belonged to genus Streptomyces and a few strains represented the rare actinobacteria Micromonospora, Rhodococcus and Tsukamurella. A total of 39 strains from G. inflate and 27 strains from G. glabra showed antimicrobial activity against at least one indicator organism. The range of the antimicrobial activity of the strains isolated from G. glabra and G. inflate was similar. A total of 34 strains from G. inflate and 29 strains from G. glabra carried at least one of the genes encoding polyketide synthases, non-ribosomal peptide synthetase and FADH2-dependent halogenase. In the type II polyketide synthase KSα gene phylogenetic analysis, the strains were divided into two major clades: one included known spore pigment production-linked KSα sequences and other sequences were linked to the production of different types of aromatic polyketide antibiotics. Based on the antimicrobial range, the isolates that carried different KSα types were not separated from each other or from the isolates that did not carry KSα. The incongruent phylogenies of 16S rRNA and KSα genes indicated that the KSα genes were possibly horizontally transferred. In all, the liquorice plants were a rich source of biocontrol agents that may produce novel bioactive compounds.
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OxyR is a key regulator in response to oxidative stress in Streptomyces avermitilis
More LessThe role of the H2O2-sensing transcriptional regulator OxyR in oxidative stress responses in Streptomyces avermitilis was investigated. An oxyR deletion mutant was more sensitive to H2O2 and tert-butyl hydroperoxide than was the WT strain, indicating that OxyR mediates the defensive system against H2O2 and organic peroxide. Evidence presented herein suggests that in cells treated with exogenous H2O2, the oxidized form of OxyR activated expression of ahpCD by binding to a palindromic sequence of the promoter region. Oxidized OxyR also induced expression of other antioxidant enzymes (KatA1, KatA2, KatA3 and OhrB1) and oxidative stress regulators (CatR, OhrR and σR). The thiol-oxidative stress regulator gene sigR was regulated at the transcription level by OxyR. We conclude that OxyR is necessary to activate transcription of sigR from the σR-dependent promoter to express an unstable larger isoform of σR during oxidative stress. In response to oxidative stress, OxyR facilitates rapid production of H2O2-scavenging enzymes to repair oxidative damage through direct regulation and cascaded regulation of CatR, OhrR and σR.
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GntR family regulator SCO6256 is involved in antibiotic production and conditionally regulates the transcription of myo-inositol catabolic genes in Streptomyces coelicolor A3(2)
More LessSCO6256 belongs to the GntR family and shows 74 % identity with SCO6974, which is the repressor of myo-inositol catabolism in Streptomyces coelicolor A3(2). Disruption of SCO6256 significantly enhanced the transcription of myo-inositol catabolic genes in R2YE medium. The purified recombinant SCO6256 directly bound to the upstream regions of SCO2727, SCO6978 and SCO6985, as well as its encoding gene. Footprinting assays demonstrated that SCO6256 bound to the same sites in the myo-inositol catabolic gene cluster as SCO6974. The expression of SCO6256 was repressed by SCO6974 in minimal medium with myo-inositol as the carbon source, but not in R2YE medium. Glutathione-S-transferase pull-down assays demonstrated that SCO6974 and SCO6256 interacted with each other; and both of the proteins controlled the transcription of myo-inositol catabolic genes in R2YE medium. These results indicated SCO6256 regulates the transcription of myo-inositol catabolic genes in coordination with SCO6974 in R2YE medium. In addition, SCO6256 negatively regulated the production of actinorhodin and calcium-dependent antibiotic via control of the transcription of actII-ORF4 and cdaR. SCO6256 bound to the upstream region of cdaR and the binding sequence was proved to be TTTCGGCACGCAGACAT, which was further confirmed through base substitution. Four putative targets (SCO2652, SCO4034, SCO4237 and SCO6377) of SCO6256 were found by screening the genome sequence of Strep. coelicolor A3(2) based on the conserved binding motif, and confirmed by transcriptional analysis and electrophoretic mobility shift assays. These results revealed that SCO6256 is involved in the regulation of myo-inositol catabolic gene transcription and antibiotic production in Strep. coelicolor A3(2).
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The enigmatic lack of glucose utilization in Streptomyces clavuligerus is due to inefficient expression of the glucose permease gene
More LessStreptomyces clavuligerus ATCC 27064 is unable to use glucose but has genes for a glucose permease (glcP) and a glucose kinase (glkA). Transformation of S. clavuligerus 27064 with the Streptomyces coelicolor glcP1 gene with its own promoter results in a strain able to grow on glucose. The glcP gene of S. clavuligerus encodes a 475 amino acid glucose permease with 12 transmembrane segments. GlcP is a functional protein when expressed from the S. coelicolor glcP1 promoter and complements two different glucose transport-negative Escherichia coli mutants. Transcription studies indicate that the glcP promoter is very weak and does not allow growth on glucose. These results suggest that S. clavuligerus initially contained a functional glucose permease gene, like most other Streptomyces species, and lost the expression of this gene by adaptation to glucose-poor habitats.
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Isolation of the biosynthetic gene cluster for tautomycetin, a linear polyketide T cell-specific immunomodulator from Streptomyces sp. CK4412
More LessThe bacterial genus Streptomyces has long been appreciated for its ability to produce various kinds of medically important secondary metabolites, such as antibiotics, anti-tumour agents, immunosuppressants and enzyme inhibitors. Tautomycetin (TMC), which is produced by Streptomyces sp. CK4412, is a novel activated T cell-specific immunosuppressive compound with an ester bond linkage between a terminal cyclic anhydride moiety and a linear polyketide chain bearing an unusual terminal alkene. Using a Streptomyces polyketide methylmalonyl-CoA acyltransferase gene as a probe, three overlapping cosmids were isolated from the genomic library of TMC-producing Streptomyces sp. CK4412. Sequence information of an approximately 70 kb contiguous DNA region revealed two multi-modular type I polyketide synthases (PKSs), and 12 additional gene products presumably involved in TMC biosynthesis. The deduced roles for most of the TMC PKS catalytic domains were consistent with the expected functions necessary for TMC chain elongation and processing. In addition, disruption of a putative TMC acyl-CoA transferase gene, located upstream of the PKS gene locus, completely abolished TMC biosynthesis. Taken together, these data provide strong supporting evidence that the cloned gene cluster identified in this study is responsible for TMC biosynthesis in Streptomyces sp. CK4412, and set the stage for detailed genetic and biochemical studies of the biosynthesis of this important metabolite.
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Inactivation of the 20S proteasome in Streptomyces lividans and its influence on the production of heterologous proteins
More LessProteasomes are self-compartmentalizing proteases first discovered in eukaryotes but also occurring in archaea and in bacteria belonging to the order Actinomycetales. In bacteria, proteasomes have so far no known function. In order to evaluate the influence of the 20S proteasome on the production of heterologous proteins by Streptomyces lividans TK24, the production of a number of heterologous proteins, including soluble human tumour necrosis factor receptor II (shuTNFRII) and salmon calcitonin (sCT), was compared with the wild-type TK24, a proteasome-deficient mutant designated PRO41 and a strain complemented for the disrupted proteasome genes (strain PRO41R). S. lividans cells lacking intact proteasome genes are phenotypically indistinguishable from the wild-type or the complemented strain containing functional proteasomes. Using the expression and secretion signals of the subtilisin inhibitor of Streptomyces venezuelae CBS762.70 (Vsi) for shuTNFRII and those of tyrosinase of Streptomyces antibioticus (MelC1) for the production of sCT, both proteins were secreted in significantly higher amounts in the strain PRO41 than in the wild-type S. lividans TK24 or the complemented strain PRO41R. However, the secretion of other heterologous proteins such as shuTNFRI was not enhanced in the proteasome-deficient strain. This suggests that S. lividans TK24 can degrade some heterologous proteins in a proteasome-dependent fashion. The proteasome-deficient strain may therefore be useful for the efficient production of these heterologous proteins.
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Evolution of the PPM-family protein phosphatases in Streptomyces: duplication of catalytic domain and lateral recruitment of additional sensory domains
More LessOriginally identified from eukaryotes, the Mg2+- or Mn2+-dependent protein phosphatases (PPMs) are a diverse group of enzymes whose members include eukaryotic PP2C and some prokaryotic serine/threonine phosphatases. In a previous study, unexpectedly large numbers of PPMs were identified in two Streptomyces genomes. In this work, a phylogenetic analysis was performed with all the PPMs available from a wide variety of microbial sources to determine the evolutionary origin of the Streptomyces PPM proteins. Consistent with earlier hypotheses, the results suggested that the microbial PPMs were relatively recent additions from eukaryotic sources. Results also indicated that the Streptomyces PPMs were divided into two major subfamilies at an early stage of their emergence in Streptomyces genomes. The first subfamily, which contains only six Streptomyces PPMs, possesses a catalytic domain whose sequence and architecture are similar to that of eukaryotic PPMs; the second subfamily contains 89 Streptomyces PPMs that lack the 5a and 5b catalytic domain motifs, similar to the PPMs SpoIIE and RsbU of Bacillus subtilis. Significant gene duplication was observed for the PPMs in the second subfamily. In addition, more than half (54 %) of the Streptomyces PPMs from the second subfamily were found to have at least one additional sensory domain, most commonly the PAS or the GAF domain. Phylogenetic analysis showed that these domains tended to be clustered according to the putative physiological functions rather than taxonomic relationship, implying that they might have arisen as a result of domain recruitment in a late evolutionary stage. This study provides an insight into how Streptomyces spp. may have expanded their PPM-based signal transduction networks to enable them to respond to a greater range of environmental changes.
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Posttranslational processing of the xylanase Xys1L from Streptomyces halstedii JM8 is carried out by secreted serine proteases
The xylanase Xys1L from Streptomyces halstedii JM8 is known to be processed extracellularly, to produce a protein of 33·7 kDa, Xys1S, that retains catalytic activity but not its cellulose-binding capacity. This paper demonstrates that at least five serine proteases isolated from Streptomyces spp. have the ability to process the xylanase Xys1L. The genes of two of these extracellular serine proteases, denominated SpB and SpC, were cloned from Streptomyces lividans 66 (a strain commonly used as a host for protein secretion), sequenced, and overexpressed in S. lividans; both purified proteases were able to process Xys1L in vitro. Three other previously reported purified Streptomyces serine proteases, SAM-P20, SAM-P26 and SAM-P45, also processed Xys1L in vitro. The involvement of serine proteases in xylanase processing-degradation in vivo was demonstrated by co-expression of the xylanase gene (xysA) and the gene encoding the serine protease inhibitor (SLPI) from S. lividans. Co-expression prevented processing and degradation of Xys1L and resulted in a threefold increase in the xylanase activity present in the culture supernatant. SpB and SpC also have the capacity to process other secreted proteins such as p40, a cellulose-binding protein from S. halstedii JM8, but do not have any clear effect on other secreted proteins such as amylase (Amy) from Streptomyces griseus and xylanase Xyl30 from Streptomyces avermitilis.
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‘Streptomyces nanchangensis’, a producer of the insecticidal polyether antibiotic nanchangmycin and the antiparasitic macrolide meilingmycin, contains multiple polyketide gene clusters
More LessSeveral independent gene clusters containing varying lengths of type I polyketide synthase genes were isolated from ‘Streptomyces nanchangensis’ NS3226, a producer of nanchangmycin and meilingmycin. The former is a polyether compound similar to dianemycin and the latter is a macrolide compound similar to milbemycin, which shares the same macrolide ring as avermectin but has different side groups. Clusters A–H spanned about 133, 132, 104, 174, 122, 54, 37 and 59 kb, respectively. Two systems were developed for functional analysis of the gene clusters by gene disruption or replacement. (1) Streptomyces phage ϕC31 and its derived vectors can infect and lysogenize this strain. (2) pSET152, an Escherichia coli plasmid with ϕC31 attP site, and pHZ1358, a Streptomyces–Escherichia coli shuttle cosmid vector, both carrying oriT from RP4, can be mobilized from E. coli into NS3226 by conjugation. pHZ1358 was shown to be generally useful for generating mutant strains by gene disruption and replacement in NS3226 as well as in several other Streptomyces strains. A region in cluster A (∼133 kb) seemed to be involved in nanchangmycin production because replacement of several DNA fragments in this region by an apramycin resistance gene [aac3(IV)] gave rise to nanchangmycin non-producing mutants.
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Physical mapping shows that the unstable oxytetracycline gene cluster of Streptomyces rimosus lies close to one end of the linear chromosome
More LessA restriction map of the 8 Mb linear chromosome of Streptomyces rimosus R6-501 was constructed for the enzymes AseI (13 fragments) and DraI (7 fragments). Linking clones for all 12 AseI sites and 5 of the 6 DraI sites were isolated. The chromosome has terminal inverted repeats of 550 kb, which are the longest yet reported for a Streptomyces species. The oxytetracycline gene cluster lies about 600 kb from one end, which might account for its frequent spontaneous amplification and deletion. Several other markers were localized on the chromosome (dnaA and recA, the rrn operons, the attachment site for pSAM2 and prophages RP2 and RP3). Comparison of the conserved markers with the map of Streptomyces coelicolor A3(2) suggested there are differences in genome organization between the two species.
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Biosynthesis of indole-3-acetic acid via the indole-3-acetamide pathway in Streptomyces spp.
More LessVarious Streptomyces spp. including S. violaceus, S. scabies, S. griseus, S. exfoliatus, S. coelicolor and S. lividans secrete indole-3-acetic acid (IAA) when fed with l-tryptophan (Trp). Production of IAA was detected in Streptomyces strains causing potato scab as well as in non-pathogenic strains. The pathways for IAA synthesis from Trp were investigated in S. violaceus and S. exfoliatus. Indole-3-acetamide (IAM), indole-3-lactic acid (ILA), indole-3-ethanol (IEt) and IAA were identified by HPLC and GC-MS. Streptomyces cells were capable of catabolizing IAM, ILA, IEt and indole-3-acetaldehyde (IAAId) into IAA. Incorporation of radioactivity into IAM, IAA and IAL but not IEt was detected when cells were fed with l-[3-14C]tryptophan. Results indicate the presence of the IAM pathway (Trp → IAM → IAA) and the possible presence of additional pathways for IAA biosynthesis in Streptomyces.
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Sporulation without aerial mycelium formation on agar medium by Streptomyces bikiniensis HH1, an A-factor-deficient mutant
More LessStreptomyces bikiniensis HH1, an A-factor-deficient mutant that did not form aerial mycelium on agar medium unless supplemented with A-factor, produced spores abundantly within colonies. The spores formed on reproductive branches morphologically similar to aerial hyphae except that they did not emerge from the surface of the colonies. The spores were morphologically more heterogeneous than those formed when A-factor was added.
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Isolation and Characterization of Actinophages Infecting Streptomyces Species and Their Interaction with Host Restriction-Modification Systems
More LessNine different phages, øA1 to øA9, were isolated from soil samples on Streptomyces antibioticus ATCC 11891, a strain which produces the macrolide antibiotic oleandomycin. Each phage displayed a different host-range which did not extend beyond Streptomyces species. Host-range was mainly limited by adsorption specificity and host-controlled restriction-modification systems. All the phages except øA3 and øA9 formed turbid plaques on S. antibioticus, but did not lysogenize this host. However, three of the phages (øA5, øA7 and øA8) were identified as temperate, since they were able to lysogenize other Streptomyces strains. All of the phages were morphologically similar and belonged to group B of Bradley’s classification. They had polyhedral heads and long, non-contractile tails. øA5, øA6 and øA7 had a base plate at the terminal end of the tail. Analysis with restriction endonucleases indicated that the nine phages contained double-strandpd DNA. Hybridization studies between the phage genomes, together with results on genome structure, allowed classification of the phages into five groups: (I) øA2, øA4 and øA9, (II) øA3 and øA8, (III) øA7, (IV) øA5 and øA6, and (V) øA1.
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Restriction Mapping and Close Relationship of the DNA of Streptomyces erythraeus Phages 121 and SE-5
More LessSUMMARY: The biological properties and genome structure of two actinophages, 121 and SE-5, infecting Streptomyces erythraeus were characterized. They had the same host range (limited to S. erythraeus) and similar DNA G + C contents (around 60 mol %). Restriction maps of their genomes also showed many similarities. The close relationship between the two phages was confirmed by DNA hybridization experiments: large parts of their genomes were homologous, except for a segment in the middle of the map, where no hybridization was detected.
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Occurrence of the Stringent Response in Streptomyces sp. and its Significance for the Initiation of Morphological and Physiological Differentiation
More LessSummary: Streptomyces sp. MA406-A-1 produced formycin (a nucleoside antibiotic) in parallel with cell growth in a synthetic medium. When the synthetic medium was supplemented with 1% (w/v) Casamino acids, however, formycin was produced only after the end of exponential growth. The intracellular ppGpp pool increased gradually towards the end of exponential growth and was maximal at the beginning of formycin production. After shift down from Casamino acids medium to synthetic medium, the ppGpp pool increased immediately, while the GTP pool decreased; under such conditions, the ability to produce formycin increased eightfold. Relaxed (rel) mutants, the first isolated for a Streptomyces species, were found at high incidence (10%) among spontaneous thiopeptin-resistant isolates and had severely reduced abilities to accumulate ppGpp. These rel mutants also failed to produce formycin under the usual culture conditions and exhibited numerous pleiotropic effects such as an inability to produce melanin and an extended delay of aerial mycelium formation. Thus Streptomyces sp. exhibited a typical stringent response, and the response initiated (or was needed for) the induction of secondary metabolism. The response may have also participated in the initiation of aerial mycelium formation by decreasing the intracellular GTP pool.
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The Loss of a Large DNA Fragment is Associated with an Aerial Mycelium Negative (Amy–) Phenotype of Streptomyces cattleya
Hybridization of various Streptomyces cattleya aerial mycelium negative (Amy–) mutants with a probe containing the gene for argininosuccinate synthetase (pTG17) has revealed the presence of two different types of mutants (stable and unstable). Stable mutants appear to have lost all or part of the region covered by the probe, while the unstable mutants demonstrate no detectable changes in this region. In one group of stable mutants (those demonstrating a partial loss of sequences hybridizing to the probe), a 4·17 kb extrachromosomal element was detected, which hybridized with the pTG 17 probe. The significance of this finding is discussed with reference to the genetic instability of the genus Streptomyces.
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Effect of Serine Hydroxamate and Methyl α-d-glucopyranoside Treatment on Nucleoside Polyphosphate Pools, RNA and Protein Accumulation in Streptomyces hygroscopicus
More LessThe accumulation of RNA and protein and the kinetics of nucleoside triphosphate and guanosine polyphosphate pools during amino acid starvation and carbon source downshift were investigated in Streptomyces hygroscopicus. RNA accumulation was controlled stringently during both amino acid starvation and carbon source downshift. The pool size of ppGpp increased dramatically under these conditions. However, the intracellular concentrations of nucleoside triphosphates were low and the concentration of guanosine polyphosphates was much lower than in Escherichia coli. The possible significance of this phenomenon in the regulation is discussed.
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A Conditional Aerial Mycelium-Negative Mutant Of Streptomyces Fradiae with Deficient Ornithine Carbamoyltransferase Activity
More LessA mutant defective in ornithine carbamoyltransferase activity and having a concomitant aerial mycelium-negative phenotype was isolated from Streptomyces fradiae. The aerial mycelium formation of the mutant could be restored by replacing l-arginine with l-citrulline in the minimal medium. The possibility that the ornithine cycle is connected with the regulation of aerial mycelium formation is discussed.
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Genetic Studies of the Fertility Plasmid SCP2 and its SCP2* Variants in Streptomyces coelicolor A3(2)
More LessThe plasmid SCP2, initially discovered through the occurrence of a high fertility variant, SCP2*, is a self-transmissible fertility factor capable of promoting chromosomal recombination within Streptomyces coelicolor A3(2). Further high fertility variants of SCP2, similar to SCP2*, were isolated from amongst the recombinants produced in matings involving SCP2, and their genetic properties were compared. SCP2 and its derivatives elicit lethal zygosis on transfer into an SCP2− recipient; this plasmid-determined phenotype allowed the isolation of SCP2− strains and the detection of the interspecific transfer of SCP2* by mating from S. coelicolor to Streptomyces parvulus and Streptomyces lividans, whereupon it underwent stable maintenance. The transfer genes of SCP2 and SCP2*, which are not normally fully expressed, were shown to undergo transient derepression on entry into an SCP2− strain. An ‘entry disadvantage’ system determined by SCP2 and SCP2* was recognized.
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